Tricyclo-DNAs (tc-DNA) are a class of constrained DNA analogs in which each nucleotide is modified by the introduction of a cyclopropane ring to restrict conformational flexibility of the backbone and to optimize the backbone geometry of the torsion angle γ. Homobasic adenine- and thymine-containing tc-DNAs form extraordinarily stable A-T base pairs with complementary RNAs.
Recently, the present inventors have proposed to use the advantageous properties of this class of nucleic acids in antisense oligonucleotides for the treatment of a number of diseases. International application No. PCT/EP2010/054735 discloses synthetic antisense oligonucleotides and methods employing antisense oligonucleotides for modifying splicing events that occur during pre-mRNA processing or for down-regulating the expression of mutated mRNA that contain repeated sequences such as, for example, 3′ or 5′ CUG, CAG, and/or CCUG. More specifically, it was shown that tricyclo-DNA antisense oligonucleotides are effective in facilitating exon skipping during pre-mRNA processing, in masking intronic silencer sequences and/or stem-loop sequences in pre-mRNA, and in targeting the RNase-mediated destruction of mRNA.
Duchenne Muscular Dystrophy (DMD) is the most common hereditary myopathy, afflicting about one in 3,500 males regardless of ethnicity. The foremost consequence of DMD is that muscle fibers become particularly fragile and natural muscle activity provokes general damage in muscle tissue. Lack of dystrophin makes that muscle fibers are particularly vulnerable to mechanical stress, and undergo recurrent cycles of necrosis. As a result, patients display progressive weakness of skeletal muscles, which are with time replaced by adipofibrotic tissue, leading to loss of ambulation around the age of twelve, whereupon premature death is caused by either respiratory failure or cardiomyopathy between the second and fourth decade. In addition, about one third of DMD patients also display cognitive impairment suggesting a noteworthy disruption of neuronal and brain function. DMD affects all voluntary muscles and involves the heart and breathing muscles in later stages of the disease. Heart and CNS should thus preferably be targeted by any therapy implemented to treat or alleviate the symptoms of DMD patients.
A new class of compounds was sought that have improved efficiency when compared to tricyclo-DNA oligonucleotides. The present invention describes the synthesis, properties and uses of tricyclo-phosphorothioate nucleotides.